Reproductive failures associated with antibodies against
caprine arthritis-encephalitis virus,
Toxoplasma gondii
and
Neospora caninum
in goats in the state of Sao Paulo, Brazil
Falhas reprodutivas associadas com a presença de do vírus da artrite-encefalite caprina,
Toxoplasma gondii e Neospora caninum em caprinos no estado de São Paulo, Brasil
Heni Falcão da COSTA1; Anne Valéria Mendonça STACHISSINI1; Hélio LANGONI1; Carlos Roberto PADOVANI2; Solange Maria GENNARI3; José Rafael MODOLO1*
1 Department of Veterinary Hygiene and Public Health, School of Veterinary Medicine and Animal Science, São Paulo State University, Botucatu-SP, Brasil
2 Department of Biostatistics, Institute of Biosciences, São Paulo State University, Botucatu-SP, Brasil
3 Department of Preventive Veterinary Medicine and Animal Health, Faculty of Veterinary Medicine and Animal Science, São Paulo-SP, Brasil
Abstract
This study aimed at assessing the occurrence of antibodies against the caprine arthritis-encephalitis virus (CAEV),
Toxoplasma gondii and Neospora caninum, as well as the associations between the presence of antibodies and the occurrence of reproductive failures in goats. Serum samples were collected from 923 goats of both sexes, over 3 months of age, from 17 dairy farms located in different municipalities of São Paulo State, Brazil. Infections by T. gondii, N. caninum and CAEV were evaluated by indirect methods of diagnosis based on indirect fluorescence antibody test (IFAT),
Neospora agglutination test (NAT), and agar gel immunodiffusion (AGID), respectively. A survey was conducted on the farms to obtain information about reproduction dates (abortions, stillbirths and births of weak and premature kids) and zoosanitary management. Antibodies against CAEV, T. gondii and N. caninum was foundin 37.81%, 23.62% and 17.23% respectively. There was no significant association between the presence of anti-CAEV antibodies and CAEV/T. gondii or CAEV/N. caninum co-infection, suggesting that CAEV does not predispose goats to infection by these agents. However, when CAEV/T. gondii (p<0.01) orCAEV/N. caninum (p<0.001) co-infection was present, the occurrence of reproductive failures was significantly higher what could indicate that CAEV-induced immunosuppression may predispose goats to develop the clinical symptoms of toxoplasmosis and neosporosis increasing the risks of the reproductive failures.
Keywords:Goat. CAEV. Toxoplasma gondii. Neospora caninum. Co-infection. Reproductive failures.
Resumo
O objetivo do presente estudo foi avaliar a ocorrência de anticorpos para o vírus da atrite-encefalite caprina (CAEV),
Toxoplasma gondii e Neospora caninum e de possíveis associações entre a presença de anticorpos e aocorrência de problemas reprodutivos em caprinos. Para tanto, foram colhidas amostras sangüíneas de 923 caprinos de ambos os sexos, acima de três meses de idade e oriundos de 17 propriedades leiteiras, de diferentes municípios do estado de São Paulo, Brasil. Os diagnósticos para T. gondii, N. caninum e CAEV foram baseados, respectivamente, na reação de imunofluorescência indireta (RIFI), teste de aglutinação para Neospora (NAT) e a imunodifusão em gel de ágar (IDGA). Um inquérito epidemiológico foi aplicado nas propriedades para obtenção de informações sobre dados reprodutivos (abortamentos, natimortalidade e nascimentos de filhotes fracos e prematuros) e de manejo zoossanitário. As ocorrências de anticorpos foram de 37,81% para CAEV, de 23,62% para T. gondii e de 17,23% para N. caninum. Não houve associação significativa entre a presença de anticorpos anti-CAEV e co-infecção com T. gondii ou N. caninum, sugerindo que o CAEV não predispõe os caprinos à infecção por estes agentes. Entretanto, quando havia, nas fazendas, animais com co-infecção pelo CAEV e T. gondii (p<0,01) ou CAEV e N. caninum (p<0,001) as ocorrências de falhas reprodutivas foram significativamente maiores, sugerindo que a imunossupressão causada pelo CAEV pode predispor os caprinos ao desenvolvimento de sintomas clínicos da toxoplasmose e neosporose, potencializando os riscos da ocorrência de problemas reprodutivos causados por estas enfermidades.
Palavras-chave: Caprino. CAEV. Toxoplasma gondii. Neospora caninum. Falhas reprodutivas.
Correspondence to:
José Rafael Modolo
Department of Veterinary Hygiene and Public Health School of Veterinary Medicine and Animal Science São Paulo State University
Distrito de Rubião Junior, s/n, P.O. box 524 CEP 18618-000, Botucatu, SP, Brasil e-mail: jrmodolo@fmvz.unesp.br
Recebido: 14/02/2010 Aprovado: 29/02/2012
Introduction
Goats infected by the caprine arthritis-encephalitis virus (CAEV) are quite predisposed to other diseases. This is partly explained by the depressive effect that may be exerted by this lentivirus on immunological reserves1. Nevertheless, associations of CAEV with
other agents remain unreported.
Toxoplasmosis commonly affects small ruminants causing reproductive problems that lead to great eco-nomic losses. Goats are one of the most susceptible species to T. gondii2,3. According to Tizard4, infections
caused by protozoa are common in immunodepressed animals. In humans with acquired immunodeficiency syndrome (AIDS), toxoplasmosis can be aggressive and frequently fatal leading to infection of the central and visceral nervous system, as well as lymph nodes5.
Studies conducted by Witt et al.6 and Lucas et al.7 in
felines infected with feline immunodeficiency virus (FIV) showed that T. gondii infection is also more fre-quent in FIV-positive animals.
Neosporosis, in turn, is known to be not so relevant in ovine and caprines, but its occurrence should also be considered as it has already been demonstrated to be associated with the birth of weak and premature kids8. Tarantino et al.9, studying a dog with
Leish-mania infantum and Neospora caninum co-infection suggested that the immunosuppressive effects of
Leishmania infection or long-term steroid therapy contributed to the development of N. caninum in this dog. Also in dogs, Cringoli et al.8 and Gennari
et al.10 observed a significantly higher occurrence of
N. caninum seropositivity among dogs that were L. infantum-positive.
The purpose of this study was to assess the associa-tions of CAEV antibodies with the presence of anti-bodies against T. gondii and N. caninum, as well as to determine whether CAEV/T. gondii or CAEV/N. caninum co-infection influence the occurrence of re-productive problems in goats.
Material and Methods
Serum samples were collected from 923 goats of both sexes, older than 3 months and originated from 17 dairy farms located in different municipalities of the State of São Paulo, Brazil. Serum samples were kept at -20ºC until analyze (agar gel immunodiffu-sion, AGID, for CAEV, indirect fluorescence antibody test, IFAT, for T.gondii and Neospora agglutination test, NAT, for N. caninum).
This study was approved by the Ethics Committee (CEEA) of School of Veterinary Medicine and Animal Science (UNESP) Botucatu, São Paulo, Brazil, with protocol number 17/2005.
During blood collection, a survey was conducted to obtain information on management and reproduc-tion dates (aborreproduc-tions, stillbirths, and weak offspring). Agar Gel Immunodiffusion (AGID) was performed according to Cultip et al.11. The antigen used was
pro-duced by the CAEV sample provided by Dr. Yahia Che-bloures of Laboratorie Associe de Recherches Sur Lês Lentivirus Chez Lês Petits Ruminants, INRA-ENVL, France, isolated by Crawfod et al.12 and replicated
with-in the caprwith-ine synovial membrane by Abreu et al.13.
Indirect Fluorescence Antibody (IFAT) was per-formed using fluoresceín-isothiacin-labeled anti-goat IgG provided by the Laboratory of Zoonoses of Botu-catu, Veterinary and Animal Science Faculty -UN-ESP together with the Center for Zoonoses Control of Municipality of São Paulo, according to Camargo14.
Samples with a titer ≥16 were considered positive15.
Anti-N. caninum antibodies were assayed by the
Neospora Agglutination Test (NAT) as described by Romand et al.16. N. caninum strain NC-1 tachyzoites
were used with a cutoff value of 1:25. This test was performed in the Animal Parasitic Diseases Labora-tory, United States Department of Agriculture, Belts-ville, USA.
The associations of CAEV with T. gondii and N. caninum were analyzed by Goodman’s association
test and the Chi-square test17,18. Associations between
agents and goat reproductive failures were assessed by the non-parametric test of Mann-Whitney19 with
sig-nificant level of 5%.
Results
The analysis of the 923 serum samples showed that 349 (37.81%) of the animals were positive for CAEV, 218 (23.62%) for T. gondii and 159 (17.23%) for N. caninum. Overall, 344 (37.27%) were positive for at least one of the agents, 143 (15.49%) for two of the agents and 32 (3.47%) for all three agents under in-vestigated (Table 1).
Table 2 shows the association of T. gondii and N. caninum distributed according to CAEV positivity or negativity. There was no significant association among
the presence of coccidian parasites in the CAEV-pos-itive or CAEV-negative goats (p>0.05).
Reproductive failures had no significant association (p >0.05%) with positivity for either T. gondii or N. caninum alone, but was significantly associated with CAEV positivity (p <0.05). However, when CAEV/T. gondii (p <0.01) or CAEV/N. caninum (p <0.001) co-infections were present, the occurrence of reproduc-tive failures was significantly higher (Table 3).
Discussion
Intercurrent infections associated with human or feline lentiviruses have been often described in the literature, probably due to the fact that these immu-nosuppressive agents lead to secondary infections caused by opportunistic parasites such as T. gondii20.
CAEV1 T. gondii2 N. caninum3 Goats
Positive % - - - 404 43.77 - - + 42 4.55 - + - 78 8.45 - + + 50 5.42 + - - 224 24.27 + - + 35 3.79 + + - 58 6.28 + + + 32 3.47
Table 1 - Frequency (%) of antibodies anti-caprine arthritis-encephalitis virus (CAEV), Toxoplasma gondii (T. gondii) and
Neospora caninum (N. caninum) in 923 goats from the state of São Paulo - Brazil
1Agar Gel Immunodiffusion 2Indirect Fluorescence Antibody 3Neospora Agglutination Test
Table 2 - Associations among the presence of antibodies anti-CAEV, T. gondii and N. caninum in 923 goats from the state of São Paulo - Brazil
T. gondii and N. caninum Associations P
CAEV T1(-) N2 (-) T(-) N (+) T(+) N (-) T(+) N (+) value
- 404 (70,38%) aA 42 (7,32%) aC 78 (13,59%) aB 50 (8,71%) aA <0.001
+ 224 (64,18%) aA 35 (10,03%) aC 58 (16,62%) aB 32 (9,17%) aA <0.001
P value >0.05 >0.05 >0.05 >0.05
Same capital letters in the line – P >0.05; same lower letters in the column – P >0.05 1Toxoplasma gondii 2Neospora caninum
However, reports on the goat lentivirus (CAEV) are still lacking.
In this study, antibodies against T. gondii or N. cani-num were similarly observed in both CAEV-positive and CAEV-negative goats. These findings do not agree with those obtained in other studies of feline lentivirus/T. gondii co-infection6,7,21 where the
fre-quency of T. gondii-seropositive cats was higher in the animals with FIV than in those without this viral infection. According to Heidel et al.20, FIV infection
induces changes in the defense mechanism that may reactivate latent infections in these animals.
The present study, no significant association was observed between positivity for T. gondii and/or N. caninum and reproductive failures such as abortion, stillbirth and weak and debilitated kids.The occur-rence of anti-T. gondii antibodies not associated with clinical problems is commonly observed in goats15,22,23.
Similarly, clinical toxoplasmosis is relatively rare in humans, occurring in 10- 20% of the cases of
infec-tion24,25. Nevertheless, symptoms of T. gondii infection
are very common in HIV-infected individuals, and toxoplasmosis is an important cause of mortality and morbidity among them26.
This is in agreement with Modolo et al.27 who found
no significant association (p>0.05) between the pres-ence of anti-CAEV antibodies and the occurrpres-ence of reproductive failures. However, when CAEV/T. gon-dii (p<0.01) or CAEV/N. caninum (p<0.001)
co-in-fections were detected in the goats, the occurrence of reproductive failures was significantly higher, indicat-ing that the viral infection could be responsible for the reactivation of infection by T. gondii or N. caninum
and clinical signs including reproductive failures. Reports of N. caninum-positivity associated with concomitant viral infections in goats are not found in the literature. Yet, studies using bovines have shown that bovine herpesvirus type 1 (BHV1) and bovine viral diarrhea (BVD) were not considereda risk fac-tor for the occurrence of N. caninum28 the same was
observed by Bartels et al.29 who also found no
associa-tion between aborassocia-tion and bacterial or viral agents. Cringoli et al.8, in Italy, reported a higher occurrence
of infection with N. caninum in dogs positive for
Leishmania infantum. In addition, Gennari et al.10,
in Araçatuba, SP, Brazil, found that the occurrence of anti-N. caninum antibodies (32.6%) was 8.6-fold higher (p≤ 0.05) in dogs naturally infected by L. cha-gasi than in those negative for this agent (3.8%). On the other hand, similar studies with both L. chagasi
positive- and negative dogs in Campo Grande, failed to demonstrate any association with seropositivity for
N. caninum30.
Our results suggest that CAEV infection may pre-dispose goats to develop the clinical symptoms of in-fection by T. gondii and N. caninum, increasing the risks of the reproductive failures that may follow these diseases.
Table 3 - Descriptive measurements of the occurrence of the infectious agents investigated according to reproductive failures - São Paulo - 2006
Infectious agent Reproductive Failure P value
Present Absent CAE 57,82a (14.67; 69.84) 20.00b (0.00; 49,15) <0.05 T. gondii 13,01a (4.76; 77.14) 6.41a (0.00; 35,90) >0.05 N. caninum 17.10a (0.00; 81.25) 5.13a (0.00; 78,21) >0.05 CAEV \ T. gondii 9.23a (0.00; 56.25) 0.00b (0.00; 6,06) <0.01 CAEV \ N caninum 7.27a (1.33; 44.29) 0.00b (0.00; 1,69) <0.001
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